Sepsis is a systemic inflammatory host response to microbial infection, with mortality of patients with severe sepsis and septic shock reaching 30% to 70%, respectively. Recombinant activated protein C (APC) is the first, and as yet only drug demonstrated to reduce mortality of patients with severe sepsis, but may cause severe bleeding, and appears to lack efficacy in children and in patients with less than severe sepsis. The mechanisms mediating the therapeutic efficacy of APC in patients, and the reasons for the limitations of APC therapy are largely unknown. The current proposal employs murine models of bacterial sepsis, mutagenesis of recombinant APC, and transgenic mouse strains to address critical aspects of APC function in the treatment of sepsis. Preliminary data suggest the working hypothesis that the anticoagulant activity of APC, on which current clinical dosing is based, may not only increase the risk of severe bleeding, but also impair the fibrin-dependent clearance of bacterial pathogens and thereby diminish the efficacy of APC therapy. In contrast, the cell signaling activity of APC via the receptors EPCR and PAR1 appears to be the predominant mechanism underlying sepsis mortality reduction by APC therapy. Accordingly, recombinant APC variants with selectively reduced anticoagulant, but normal signaling activity exhibit a striking gain of efficacy in murine models of sepsis, as compared to normal APC. Proposed experiments will (AIM 1) demonstrate the harmful effects of APC's anticoagulant activity on the antibacterial host defense and reveal the underlying molecular mechanism;and (AIM 2) pinpoint the critical receptors and cellular components targeted by APC's signaling activity to reduce sepsis mortality, and (AIM 3) provide first evidence whether aggressive dosing of recombinant APC variants with selectively reduced anticoagulant activity may be expected to improve the overall efficacy and safety of APC therapy. PUBLIC HEALTH RELEVANCE: Severe sepsis after microbial infection affects in excess of 700,000 patients annually in the US alone, and is responsible for ~9% of deaths from all causes. Recombinant activated protein C (APC) is the as yet only available sepsis drug shown to reduce (by ~6%) absolute mortality of patients with severe sepsis, but may cause severe bleeding, and lacks efficacy in children and in patients with less than severe sepsis. We will investigate whether novel APC variants that selectively engage the critical molecular and cellular targets to promote survival, but lack the harmful side effects of normal APC can significantly improve the efficacy of APC therapy.